A number of studies using pH-sensitive MRI contrast agents, microelectrodes, and MR spectroscopy with hyperpolarized C13 have consistently demonstrated that the extracellular pH (pHe) of tumors is significantly lower (6.6-7.0) than healthy tissues (7.2-7.4) [Gillies R J, et al., pH imaging. A review of pH measurement methods and applications in cancers. IEEE Eng Med Biol Mag 2004; 23(5):57-64; Gillies R J, et al., MRI of the tumor microenvironment. J Magn Reson Imaging 2002; 16(4):430-50; Helmlinger G, et al., Interstitial pH and pO2 gradients in solid tumors in vivo: high-resolution measurements reveal a lack of correlation. Nat. Med. 1997; 3(2):177-82; Gallagher F A, et al., Magnetic resonance imaging of pH in vivo using hyperpolarized 13C-labelled bicarbonate. Nature 2008; 453(7197):940-3]. This acidity is primarily due to (a) anaerobic glycolysis in tumor regions subjected to short-term or long-term hypoxia as a result of poorly organized vasculature with diminished chaotic blood flow, and (b) aerobic glycolysis (the Warburg effect), a common cancer phenotypic property in which the glycolytic metabolic pathways are used even in the presence of oxygen [Gatenby R A, et al., Why do cancers have high aerobic glycolysis? Nat Rev Cancer 2004; 4(11):891-9].
An acidic pHe induces pleiotropic changes in tumor cells. In many tumor types, acute or chronic incubation in a low pH microenvironment increases invasiveness both in vitro and in vivo [Moellering R E, et al., Acid treatment of melanoma cells selects for invasive phenotypes. Clinical & experimental metastasis 2008; 25 (4):411-25]. Lowering culture pH to 6.7 has been demonstrated to result in a 4-fold increase in the number of in vivo metastases of the treated cells compared with controls after tail vein injection [Rofstad E K, et al., Acidic extracellular pH promotes experimental metastasis of human melanoma cells in athymic nude mice. Cancer Res 2006; 66(13):6699-707; Cuvier C, et al., Exposure to hypoxia, glucose starvation and acidosis: effect on invasive capacity of murine tumor cells and correlation with cathepsin (L+B) secretion. Clinical & experimental metastasis 1997; 15(1):19-25; Kalliomaki T, et al., Effects of tumour acidification with glucose+MIBG on the spontaneous metastatic potential of two murine cell lines. Brit J Cancer 2004; 90(9):1842-9]. In addition, a variety of cancer cell populations, when exposed to an acidic environment, have been shown to increase expression of interleukin-8 (IL-8), vascular-endothelial growth factor (VEGF), carboninc anhydrase IX (CAIX), lactate dehyrodgenase (LDH), cathepsin B, and matrix metalloproteinases (MMP)-2 and MMP-9, all of which are associated with increased tumor growth and invasion in-vivo [Rozhin J, et al., Pericellular pH affects distribution and secretion of cathepsin B in malignant cells. Cancer Res 1994; 54(24):6517-25; Xu L, et al., Acidic pH-induced elevation in interleukin 8 expression by human ovarian carcinoma cells. Cancer Res 2000; 60(16):4610-6; Shi Q, et al., Regulation of vascular endothelial growth factor expression by acidosis in human cancer cells. Oncogene 2001; 20(28):3751-6; Swietach P, et al., Regulation of tumor pH and the role of carbonic anhydrase 9. Cancer metastasis reviews 2007; 26(2):299-310]. Interestingly, tumor cells are typically able to maintain high proliferation rates even in an acidic environment [Ceccarini C, et al., pH as a determinant of cellular growth and contact inhibition. PNAS 1971; 68(1):229-33].
An acidic pHe, on the other hand, induces significant toxicity in normal cells by reducing proliferation [Id.] and promoting apoptosis via a p53-dependent pathway [Park H J, et al., Acidic environment causes apoptosis by increasing caspase activity. Brit J Cancer 1999; 80(12):1892-7] initiated by increasing caspase activity [Williams A C, et al., An acidic environment leads to p53 dependent induction of apoptosis in human adenoma and carcinoma cell lines: implications for clonal selection during colorectal carcinogenesis. Oncogene 1999; 18(21):3199-204]. In addition, an acidic pHe in normal tissues increases degradation of the extracellular matrix due to the production and release of proteolytic enzymes [Rozhin J, et al., Cancer Res 1994; 54(24):6517-25], promotes angiogenesis through release of VEGF [Shi Q, et al., Oncogene 2001; 20(28):3751-6], and limits immune response to tumor antigens [Lardner A. The effects of extracellular pH on immune function. J Leukocyte Biol 2001; 69(4):522-30].
These findings have been synthesized into the acid-mediated tumor invasion model, which proposes that intratumoral acidosis results in the flow of H+ ions along concentration gradients into normal tissue adjacent to the tumor. This produces a peritumoral ring of dead and dying cells and a degraded extracellular matrix into which the still viable malignant cells invade [Gatenby R A, et al., A reaction-diffusion model of cancer invasion. Cancer Res 1996; 56(24):5745-53; Gatenby R A, et al., Acid-mediated tumor invasion: a multidisciplinary study. Cancer Res 2006; 66(10):5216-23]. The model is supported by experimental evidence demonstrating a peritumoral acid gradient associated with normal cell apoptosis and extracellular matrix degradation.
Indirect support for this model is seen in a number of clinical studies, including (a) observations that increased glucose uptake on [18F]fluorodeoxyglucose positron emission tomography scans (and, therefore, increased acid production) in the transition from in situ to invasive cancer [Yasuda S, et al., 18F-FDG PET detection of colonic adenomas. J Nucl Med 2001; 42(7):989-92; Abbey C K, et al., In vivo positron-emission tomography imaging of progression and transformation in a mouse model of mammary neoplasia. PNAS 2004; 101(31):11438-43] and that a higher level of uptake in many cancer types confers poor prognosis [Schwarzbach M H, et al., Prognostic significance of preoperative [18-F] fluorodeoxyglucose (FDG) positron emission tomography (PET) imaging in patients with resectable soft tissue sarcomas. Ann Surg 2005; 241(2):286-94; Schwartz D L, et al., FDG-PET prediction of head and neck squamous cell cancer outcomes. Arch Otolaryngol 2004; 130(12):1361-7; Vansteenkiste J, et al., Positron-emission tomography in prognostic and therapeutic assessment of lung cancer: systematic review. Lancet Oncol 2004; 5(9):531-40], (b) increased intratumoral lactate concentrations is associated with a poor prognosis [Walenta S, et al., High lactate levels predict likelihood of metastases, tumor recurrence, and restricted patient survival in human cervical cancers. Cancer Res 2000; 60(4):916-21; Schwickert G, et al., Correlation of high lactate levels in human cervical cancer with incidence of metastasis. Cancer Res 1995; 55(21):4757-9], and (c) increased expression of proteins that are upregulated by acidic pHe, including IL-8, cathepsin B, lactate dehydrogenase, and carbonic anhydrase IX [Rozhin J, et al., Cancer Res 1994; 54(24):6517-25; Xu L, et al., Cancer Res 2000; 60(16):4610-6; Shi Q, et al., Oncogene 2001; 20(28):3751-6; Swietach P, et al., Cancer metastasis reviews 2007; 26(2):299-310; Ceccarini C, et al., PNAS 1971; 68(1):229-33] are associated with poor prognosis [Kolev Y, et al., Lactate dehydrogenase-5 (LDH-5) expression in human gastric cancer: association with hypoxia-inducible factor (HIF-1alpha) pathway, angiogenic factors production and poor prognosis. Ann Surg Oncol 2008; 15(8):2336-44; Hui E P, et al., Coexpression of hypoxia-inducible factors 1 alpha and 2alpha, carbonic anhydrase IX, and vascular endothelial growth factor in nasopharyngeal carcinoma and relationship to survival. Clin Cancer Res 2002; 8(8):2595-604; Choi S W, et al., Expression of carbonic anhydrase IX is associated with postoperative recurrence and poor prognosis in surgically treated oral squamous cell carcinoma. Hum Pathol 2008; 39(9):1317-22; Nomura T, et al., Involvement of cathepsins in the invasion, metastasis and proliferation of cancer cells. J Med Invest 2005; 52(1-2):1-9; Benoy I H, et al., Increased serum interleukin-8 in patients with early and metastatic breast cancer correlates with early dissemination and survival. Clin Cancer Res 2004; 10(20:7157-62].